基于受体后信号转导通路探讨疏肝理气调神法促进恶劣心境大鼠海马神经细胞重塑机制的研究

The dysthymia affective disorder is a chronic depressive state of mind as the main features of the disease, social dysfunction and disability caused by more than high blood pressure, diabetes and other diseases. 《danxixinfa》said "Once suffering from depression, suffering from other diseases". But few studies of the pathogenesis of the disease and medical treatment mechanism, it is difficult to get recognized achievements. Our previous work has shown that dispersing the stagnated liver-qi and regulating vitality can regulate the disease rat neurotransmitters and their receptors, brain-derived neurotrophic factor, HPA axis, to improve the pathomorphology in hippocampus, and to improve the symptoms of depression in rats . To further clarify the effect target of Post-receptor signal transduction pathway, we research the AC-cAMP-CREB-BDNF and HPA axis-Glu-of NMDAR-NO signal transduction pathway using RT-PCR and immunohistochemistry methods .We investigate the disease rat neurotransmitter receptors, G proteins, the HPA axis, glutamic acid and its coupling of signal transduction pathways, gene transcription, protein expression, and their mutual relations. So we can clarify mechanism from the molecular level description of the method to promote the hippocampus nerve cells to treat dysthymia to multi-level, multi-target mechanism of action, determine new biological indicators as well as disease diagnosis and criteria for assessment of therapeutic effects, and provide theoretical support for basic research.

恶劣心境是以慢性抑郁性心境为主要特征的情感障碍性疾病，所造成的社会功能障碍和残疾超过高血压、糖尿病等病。《丹溪心法》说："一有拂郁，诸病生焉"。但目前对本病发病机理和中医药治疗机制研究较少，难以取得公认成果。我们前期工作显示中医疏肝理气调神法可以调节本病大鼠单胺类神经递质及其受体、脑源性神经营养因子、HPA轴、改善海马病理形态学，并改善大鼠抑郁症状。为进一步明确该治法的作用靶点，我们以受体后AC-cAMP-CREB-BDNF和HPA轴-Glu-NMDAR-NO信号转导通路为研究内容，应用RT-PCR、免疫组化等方法探讨本病大鼠神经递质、受体、G蛋白、HPA轴、谷氨酸及其偶联的信号传导通路、基因转录、蛋白表达，及其相互联系，从分子水平说明本法重塑海马神经细胞从而治疗恶劣心境的机理，阐明本法治疗本病多层次、多靶点的作用机制，同时为该病的诊断、疗效判定提供新生物学指标，为其基础研究提供理论支撑。

本研究采用慢性不可预见的温和性应激方法，结合孤养的方法在国内领先的制备出病证统一的恶劣心境肝气郁结证大鼠动物模型；应用PT-PCR、免疫组化和光镜、电镜等手段。研究发现（1）该药物可以改善恶劣心境大鼠的行为学；（2）提高海马cAMP含量,提高海马PKA、CREB阳性细胞数，疗效与氟西汀相当；可以提高BDNF、TrkB阳性细胞数，疗效优于氟西汀；（3）可以上调BDNFmRNA水平；（4）可以改变大鼠海马C1区组织病理变化：治疗组治疗后光镜下神经元数目较模型组明显增多，固缩和空泡化现象少；电镜下观察与正常组类似，与治疗前相比明显改善。上述研究结果证实克郁舒神颗粒通过调节AC-cAMP-CREB-BDNF信号转导治疗本病的多层次、多靶点作用机制，为中医药治疗恶劣心境提供了基础理论支持。